Abstract

The phylogenetic systematics of bovin species forms a common basis for studies at
multiple scales, from the level of domestication in populations to major cladogenesis.
The main big-picture accomplishments of this productive field, including two recent
works, one in BMC Genomics, are reviewed with an eye for some of the limitations and challenges impeding progress.
See Research article http://www.biomedcentral.com/1471-2164/10/177

Commentary

Recent years have seen a flurry of interest and inquiry into the evolutionary history
of Bovini, the clade comprising living oxen (genus Bos) and the living buffaloes (genera Syncerus and Bubalus). A disproportionate interest in the phylogenetics of Bovini, relative to other bovid
or ruminant clades, no doubt stems from the importance of bovins as a prime source
of human sustenance since at least the Pleistocene. In addition, bovins are widespread
(naturally occurring on four continents), ecologically differentiated with wide habitat
tolerances (Figure 1), taxonomically diverse (around a dozen living species and over 50 fossil), and,
given their large size and affinity for wet habitats, possess a high potential for
preservation in the fossil record. For these reasons, the evolutionary record of Bovini
provides an exemplary resource for studies on evolutionary patterns and processes.

Figure 1.Bovins stand apart from other antelopes (Bovidae) in the wide range of environments
they inhabit, from high montane to wet tropical. Bovini today comprises 12 species found on four continents. (Yak: iStockphoto.com/kodda; African buffaloes: iStockphoto.com/dawnn).

One avenue of research that has been continuously providing new information, from
Miocene higher clade origins to Holocene population dynamics, has been the analysis
of bovin genomes. Deeper and more extensive mining of the genomes of bovin species
has seen a consistently improving phylogeny for this clade. A recent study by MacEachern
et al. [1] used 84 autosomal gene sequences from 15 different genes to examine phylogenetic
relationships among bovin species and populations, reconstructing geographic divergences
and intricate histories of genetic introgression and geographic divergence. Similarly,
Decker et al. [2] used a much expanded genomic data set to investigate phylogenetic relationships from
the level of Ruminantia (ruminants, including cattle, antelope, deer, giraffe, and
chevrotains) to that of domesticated cattle breeds, including DNA from the extinct
Bison priscus. Advances in DNA sequencing techniques have even produced a complete mitochondrial
genome from the aurochs (Bos primigenius) [3], the progenitor of domestic cattle (Bos taurus).

Studies such as these highlight the current focus on unraveling the history of bovin
evolution by way of the bovin genome, and the present interest in tracing the history
and geography of domestication events. However, two aspects that have seen little
progress in the last few years are the phylogenetic assembly of the total bovin clade
and the dating of major cladogenetic events within Bovini. Even in the cutting edge
studies cited above, little advance is made at the level of major bovin cladogenesis
over previous work almost a decade older. From this perspective, the literature of
the last years has provided mostly incomplete phylogenies of Bovini dated with inadequate
molecular clock estimates. This stems from several factors. First has been the dearth
of phylogenetic work on fossil bovin taxa. Second is the regular omission of certain
crucial bovin taxa from phylogenetic work. Third is a lack of precision in the phylogenetic
terminology used to communicate between paleontological and molecular studies, resulting
in the choice of poor references for molecular clock calibration.

We here present our views to highlight some significant gaps and challenges that remain
in the field of bovin phylogenetics. Our recommendations also aim to increase the
utility of studies for workers of different methodological backgrounds.

Stability in bovin systematics: mission accomplished?

A key achievement of the past two decades of molecular phylogenetic analyses has been
recurrent and consistent support for a systematic classification of Bovidae (Figure
2; and see [4]). All living bovids may be divided among either Bovinae or Antilopinae. Bovinae is
the clade uniting Bovini (buffaloes and oxen), Tragelaphini (spiral-horned antelopes,
including kudu), and Boselaphini (nilgai and chousinga). This classification requires
that members of Bovinae be referred to as bovines, while members of Bovini are bovins,
though most of the literature still uses the term bovine in reference to the Bovini.
Living bovins are further divided among the Bovina (genus Bos, including Bison) and the Bubalina (Syncerus and Bubalus). A certain amount of confusion surrounding the taxonomy of domestic derivates of
wild bovin species has also been addressed by a ruling by the International Commission
on Zoological Nomenclature (see [5]).

Figure 2.Phylogeny showing the position of Bovini in Bovidae and Bovinae, the division of Bovini
into Bovina and Bubalina, and the uncertain placement of the saola (Pseudoryx nghetinhensis).

One major remaining conundrum concerns the saola (Pseudoryx nghetinhensis), a little-understood forest antelope from Vietnam and Laos only made known to science
since 1993. This species is challenging the definition, composition, and diagnosis
of parts of the bovin phylogenetic tree. The saola's small size, simple non-divergent
horns, large preorbital fossa (bony depression anterior to the orbital cavity), and
simple-shaped teeth make it a very primitive-looking bovid, especially with respect
to the large and derived Bovini. It is therefore puzzling that molecular phylogenetic
analyses consistently place the saola within Bovini [6,7]. The exact position and relationship of the saola to the other bovin species is in
need of further confirmatory work. Knowing the relationship of this goat-like ungulate
to the remainder of Bovini is important, not just to appease curiosity about an enigmatic
forest antelope, but because the phylogenetic position of this creature may in fact
upset some of the 'stabilized' topology shown in Figure 2 (see also below).

Dating phylogenies: putting the cow before the cart

Phylogenetic studies of fossil Bovini are a crucially missing basis for bovin molecular
clock calibrations. Molecular phylogenies are being calibrated using fossil data that
are wrong or, at best, highly speculative. For example, reference calibration nodes
used in the literature include: the first appearance of Bovidae, the divergence of
Bison and Bos, the divergence of Bovina from Bubalina, and the divergence of Bovini from Tragelaphini.
However, the reality is that none of the ages of these divergence events is at all
well established (Geraads's 1992 work [8] might be the only extensive phylogenetic analysis of fossil and living bovins to
date). The phylogenetic relationships of many fossil Bovini, and, as a result, the
evolutionary history of many living Bovini, await more thorough analysis of fossil
bovin taxa.

A related issue concerning the use of fossils to calibrate molecular phylogenies stems
from an inattention to the distinction between dating crown clades and total clades
[9]. A crown clade is one defined on the basis of extant taxa, while a total clade includes
the crown plus any extinct taxa located on the ancestral 'stem' of the crown clade
(Figure 3). Despite much progress towards stability in clade names, there is still a certain
fog of confusion, much of it gone unnoticed, about clade definitions and compositions.
For example, a late Miocene fossil is assigned to 'Bovini' in the paleontological
literature without determining whether it actually belongs to the stem group or the
crown clade. In a later study, this same taxon is then simply assumed to belong in
the crown clade, and is used to date the node of origination of crown Bovini in a
molecular phylogeny. The fact that the referenced fossil taxon is just as likely to
be a stem bovin means the clade origination estimate might actually be off by millions
of years, a wide margin when considering Neogene taxa (Figure 3). The use of poorly-placed fossils to date a molecular phylogeny can only be expected
to produce spurious results. Studies seeking to assess the timing of evolutionary
events with respect to environmental changes, for example, cannot rely on such results,
nor could phylogeographic reconstructions.

Figure 3.Fossil taxa relied upon for molecular clock calibrations may produce highly spurious
dates if their phylogenetic position is not precisely known. In this example, an early fossil taxon attributed simply to 'Bovini' might be assumed
to be close to the most recent common ancestor of the living bovins (point 1, node
of origin of crown Bovini as defined by the most recent common ancestor of Bos taurus and Bubalus bubalis) when in fact it may be a stem bovin of much older age (point 2).

Until better paleontological studies are available, caution and wide margins of error
are advised when referring to the ages and phylogenetic placement of fossil bovins
for molecular clock calibration. Though commendable efforts have been made in this
regard, dates produced from molecular clock estimates will remain very imprecise so
long as the fossils used to calibrate these rates are themselves poorly understood
phylogenetically. The fossil record of Bovini is already very large and holds great
promise for dating the major cladogenetic events within Bovini, so there is every
reason for rapid progress in this regard.

From genes to genera: reconciling scales of analysis

The phylogeny of Bovini is being approached from the scale of genes to genera. Studies
of the genomes and studies of the fossil record proceed from different methodologies
and at different scales, fossils providing a relatively coarse but deep temporal perspective,
and molecular work providing a narrow but very highly resolved picture of the modern.
Both fields share a common goal, but too often the results of each approach are difficult
to reconcile with the other. One limitation to the synthesis of results is the use
of limited taxonomic sampling in a phylogenetic analysis. Limited taxonomic representation
decreases the reliability and precision of a phylogeny, in turn limiting interpretations
on dating, biogeography, and cladogenesis.

Paleontological studies should consider living taxa to the greatest extent they can.
This is important considering that workers seeking to calibrate molecular phylogenies
are ploughing the literature for information on the timing of origination of crown
(that is, extant) clades. Likewise, phylogenetic studies treating living taxa are
most informative when they too consider the largest available sample of species. For
example, paleontologists investigating the split of Bovina and Bubalina will want
to refer to phylogenies that include the entirety of living Bovini to be sure that
the node defining the crown clade has been defined to the finest degree possible.
Additionally, taxa that are rare or poorly understood have the greatest potential
to disrupt 'stable' topologies and alter previous notions, and yet such 'enigmatic'
taxa are regularly missing from analyses. An investigation seeking to unravel the
relationships among the different clades of Ruminantia cannot afford to omit the Moschidae
(musk deer, small ruminants lacking antlers and today restricted to central and northeastern
Asia). Any analysis of the Bovini cannot now afford to exclude the saola. It is precisely
the fact that such taxa are so disparate in form and restricted in distribution that
makes them most interesting for evolutionary reconstruction.

Great progress has been made towards stable systematic classifications in recent years
by workers in different fields utilizing different approaches. Continued progress
requires an effort to better integrate the different results of the geneticists, ecologists,
archaeologists, and paleontologists working on the origins of bovins. Improved communication
among workers in different fields will greatly promote the output, precision, and
accuracy of results in studies to come.

Acknowledgements

FB is supported by a National Science Foundation International Research Fellowship
Award (#0852975). This work was also supported by the Agence Nationale de la Recherche
ANR-09-BLAN-0238.

Gatesy J, Arctander P: Hidden morphological support for the phylogenetic placement of Pseudoryx nghetinhensis with bovine bovids: A combined analysis of gross anatomical evidence and DNA sequences
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